A Simple, Cost-Effective, and Extraction-Free Molecular Diagnostic Test for Sickle Cell Disease Using a Noninvasive Buccal Swab Specimen for a Limited-Resource Setting
Priya Thakur,
Pragya Gupta,
Nupur Bhargava,
Rajat Soni,
Narendra Varma Gottumukkala,
Sangam Giri Goswami,
Gaurav Kharya,
Vinodh Saravanakumar,
Padma Gunda,
Suman Jain,
Jasmita Dass,
Mukul Aggarwal,
Sivaprakash Ramalingam
Affiliations
Priya Thakur
CSIR—Institute Genomics and Integrative Biology, Mathura Road, Sukhdev Vihar, New Delhi 110025, India
Pragya Gupta
CSIR—Institute Genomics and Integrative Biology, Mathura Road, Sukhdev Vihar, New Delhi 110025, India
Nupur Bhargava
CSIR—Institute Genomics and Integrative Biology, Mathura Road, Sukhdev Vihar, New Delhi 110025, India
Rajat Soni
CSIR—Institute Genomics and Integrative Biology, Mathura Road, Sukhdev Vihar, New Delhi 110025, India
Narendra Varma Gottumukkala
CSIR—Institute Genomics and Integrative Biology, Mathura Road, Sukhdev Vihar, New Delhi 110025, India
Sangam Giri Goswami
CSIR—Institute Genomics and Integrative Biology, Mathura Road, Sukhdev Vihar, New Delhi 110025, India
Gaurav Kharya
Indraprastha Apollo Hospitals, Sarita Vihar, New Delhi 110020, India
Vinodh Saravanakumar
CSIR—Institute Genomics and Integrative Biology, Mathura Road, Sukhdev Vihar, New Delhi 110025, India
Padma Gunda
Thalassemia and Sickle Cell Society, Rajendra Nagar, Hyderabad 500052, India
Suman Jain
Thalassemia and Sickle Cell Society, Rajendra Nagar, Hyderabad 500052, India
Jasmita Dass
Department of Hematology, All India Institute of Medical Sciences, New Delhi 110029, India
Mukul Aggarwal
Department of Hematology, All India Institute of Medical Sciences, New Delhi 110029, India
Sivaprakash Ramalingam
CSIR—Institute Genomics and Integrative Biology, Mathura Road, Sukhdev Vihar, New Delhi 110025, India
Sickle cell disease (SCD) is the most prevalent life-threatening blood monogenic disorder. Currently, there is no cure available, apart from bone marrow transplantation. Early and efficient diagnosis of SCD is key to disease management, which would make considerable strides in alleviating morbidity and reducing mortality. However, the cost and complexity of diagnostic procedures, such as the Sanger sequencing method, impede the early detection of SCD in a resource-limited setting. To address this, the current study demonstrates a simple and efficient proof-of-concept assay for the detection of patients and carriers using extraction-free non-invasive buccal swab samples by isothermal DNA Amplification coupled Restrictase-mediated cleavage (iDAR). This study is a first of its kind reporting the use of buccal swab specimens for iDA in molecular diagnosis of a genetic disease, all the while being cost effective and time saving, with the total assay time of around 150 min at a cost of USD 5. Further, iDAR demonstrates 91.5% sensitivity and 100% specificity for detecting all three alleles: SS, AS, and AA, having a 100% concordance with Sanger sequencing. The applicability of the iDAR assay is further demonstrated with its adaptation to a one-pot reaction format, which simplifies the assay system. Overall, iDAR is a simple, cost-effective, precise, and non-invasive assay for SCD screening, with the potential for use in a limited resource setting.